Na+-Ca2+ exchange in inside-out cardiac sarcolemmal vesicles.

We have measured Na+-CaZ+ exchange in the insideout vesicles of highly purified cardiac sarcolemma from dog ventricles. This was accomplished in a mixed population of Sarcolemmal vesicles by first loading the inside-out vesicles with Na+ through the action of the glycoside-sensitive, ATP-dependent Na' pump. Due to the asymmetric nature of active Na' transport, this will only occur in inside-out vesicles. We then examine the exchange of extravesicular Ca2+ for the digitoxigeninsensitive component of internal Na'. We confirm that the Na+-Ca2+ exchange mechanism and the ATP-dependent Na+ pump are present on the same vesicles and that Na+-Ca2+ exchange has a sarcolemmal origin. Na+-Ca2+ exchange of inside-out Sarcolemmal vesicles is compared with Na+-Ca2+ exchange in the total population of Sarcolemmal vesicles. This is measured as the exchange of external Ca2+ for Na' taken up by passive diffusion (which will occur in both inside-out and right side-out vesicles). Evidence is presented which implies that Na+-Ca2+ exchange in the total population is due primarily to right side-out Sarcolemmal vesicles. We find that Nai+-dependent Ca2+ uptake in the inside-out vesicles and the total population of vesicles has similar values for K&!aa+), identical responses to altered pH, and similar responses to valinomycin-induced membrane potentials. These data strongly imply that the characteristics of the Ca2+ binding sites on the two sides of the exchange transport mechanism are symmetrical. Na,+-dependent Ca2+ efflux experiments suggest that the Na' binding sites of the Na+-CaZ+ exchange mechanism may be different on the opposite sides of the Sarcolemmal membrane. The dependence of Ca2+ efflux on p a + ] in inside-out vesicles implies that in vivo a small shift in intracellular Na+ levels will have marked effects on Ca2+ influx.